Agonist-dependent phosphorylation of the formyl peptide receptor is regulated by the membrane proximal region of the cytoplasmic tail.
Formyl peptide receptor (FPR) is a chemoattractant G protein-coupled receptor (GPCR) involved in the innate immune response against bacteria. Receptor activation is terminated by receptor phosphorylation of two serine- and threonine-rich regions located in the distal half of the cytoplasmic tail. In this study we show that introduction of an amino acid with a bulky side chain (leucine or glutamine) adjacent to a single leucine, L320, in the membrane-proximal half of the cytoplasmic tail, significantly enhanced receptor phosphorylation, beta-arrestin1/2 translocation, and receptor endocytosis, without affecting G(i)-mediated ERK1/2 activation and release of intracellular calcium. In addition, the point mutations resulted in diminished susceptibility to trypsin, suggesting a conformation different from that of wild type FPR. Alignment of the FPR sequence with the rhodopsin sequence showed that L320 resides immediately C-terminal of an amphipathic region that in rhodopsin forms helix 8. Deletion of seven amino acids (Delta309-315) from the predicted helix 8 of FPR (G307-S319) caused reduced cell signaling as well as defects in receptor phosphorylation, beta-arrestin1/2 translocation and endocytosis. Thus, the amino acid content in the N-terminal half of the cytoplasmic tail influences the structure and desensitization of FPR.
Suvorova
Elena S
ES
Department of Microbiology, Montana State University, 109 Lewis Hall, Bozeman, MT 59717, USA.
Gripentrog
Jeannie M
JM
Jesaitis
Algirdas J
AJ
Miettinen
Heini M
HM
D000595:Amino Acid Sequence; D000818:Animals; D019390:Arrestins / Q000378:metabolism; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D004705:Endocytosis; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D007930:Leucine / Q000378:metabolism; D008024:Ligands; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D011485:Protein Binding; D021381:Protein Transport; D044042:Receptors, Formyl Peptide / Q000819:agonists / Q000737:chemistry* / Q000378:metabolism*; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014357:Trypsin / Q000378:metabolism; D000071557:beta-Arrestins
D019390:Arrestins; D008024:Ligands; D050505:Mutant Proteins; D044042:Receptors, Formyl Peptide; D000071557:beta-Arrestins; D014357:Trypsin; D019204:GTP-Binding Proteins; D007930:Leucine